A single mutation at lysine 241 alters expression and trafficking of the D2 dopamine receptor.
Ubiquitination of G protein-coupled receptors has been identified to regulate receptor signal transduction including agonist-induced internalization and sorting of internalized receptor for degradation or for recycling. Using co-immunoprecipitation and immunoblot analysis, I found that the membrane-associated D(2) dopamine receptor (DAR) is mono-ubiquitinated in the absence of an agonist following heterologous expression in human embryonic kidney cells (HEK293). By using site-directed mutagenesis, this report shows that the loss of lysine-241, K241A D(2) DAR reduced the amount of membrane-associated D(2) DAR. It is of interest that the K241A D(2) DAR also had a distinctly different ubiquitination pattern than the wild-type D(2) DAR. It is important to note that the ubiquitinated mutant D(2) DAR was degraded through ubiquitin-proteasome pathway. These data provide the factual evidence that a loss of lysine-241 of the D(2) DAR affects receptor ubiquitination and renders the protein susceptible to the proteasomal degradation.
Kim
Ok-Jin
OJ
Department of Pharmacology and Toxicology, School of Pharmacy, University of Kansas, Lawrence, Kansas 66045-7582, USA. stevenleonardparker@msn.com
D000818:Animals; D002460:Cell Line; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D008239:Lysine / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000378:metabolism*; D021381:Protein Transport / Q000235:genetics; D051381:Rats; D017448:Receptors, Dopamine D2 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection; D025801:Ubiquitin / Q000187:drug effects / Q000378:metabolism*; D054875:Ubiquitination
D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D007976:Leupeptins; D017448:Receptors, Dopamine D2; D025801:Ubiquitin; D046988:Proteasome Endopeptidase Complex; D008239:Lysine; D005033:Ethylmaleimide; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde